Immortalised cell lines, such as the HMC3 microglia cell line, are widely used in research and drug discovery screens due to their ease of culture, scalability and low cost. It is accepted that immortalised cell lines do not best recapitulate the complexities, functions and properties of human cell types, however, what does that look like in practice?    

Here, we showcase a range of functional and phenotypical data in which our human iPSC-derived ioMicroglia demonstrate limitations in the HMC3 cell line, one which is still widely used today by both academic and industry researchers. Our investigations reveal that HMC3 cells do not express the expected protein markers, possess a transcriptomic profile significantly different from primary microglia and human iPSC-derived ioMicroglia, and display very weak or no phagocytic activity and cytokine secretion. 

This case study demonstrates the importance of complementary secondary screens in biologically relevant models closely mimicking human biology, such as human iPSC-derived ioMicroglia. Incorporating such cells offers a powerful opportunity to de-risk the drug discovery pipeline by filtering out false positives, validating on-target effects, and prioritising hits with better translational potential. Confirming candidates in an in vitro model that closely matches human microglia biology could help prevent failure at more costly steps and build confidence for downstream decisions. 
Better awareness of the limitations of immortalised cell lines will support neurodegeneration research and drug discovery, where greater fidelity is critical in translating findings.